PROJECT SUMMARY/ABSTRACT: PROJECT 3 A major hurdle to identifying disease mechanisms in amyotrophic lateral sclerosis (ALS) is the heterogeneous nature of the disease. People with ALS exhibit a wide variety of clinical presentations, rates of progression and underlying genetic risk factors. ALS is more likely a syndrome rather than a single disease that leads to a recognizable clinical phenotype that we call ALS. Our approach to unraveling the pathophysiology of ALS is to focus on a specific genetically-defined population where the disease is defined by a common underlying mechanism. Here, people carrying the C9orf72 G4C2 hexanucleotide repeat expansion mutation (c9ALS) are the genetically defined population. As opposed to sporadic ALS (sALS), where an array of genetic, environmental, and unknown factors drive disease, a common disease-causing pathophysiology underlies c9ALS. The proposition is that investigating this relatively homogenous ALS genetic subgroup will provide a window of opportunity for understanding disease mechanisms and developing treatments. The hope is that discoveries in this smaller group of patients will be applicable to the larger ALS population. In this project, we will use next generation mass spectroscopy and systems biology analysis to discover biomarkers of disease pathways and progression. In Specific Aim 1, we will interrogate human motor cortex from c9ALS and sALS patients to generate proteomic networks and modules that define c9ALS. In Specific Aim 2, we will partner with the investigators of Project 1 to define proteomic networks in a C9orf72 G4C2 mouse model; this will allow us to analyze proteomic changes in brain at several time points during disease progression. In addition, proteomic analysis of brains from mice treated with an antisense oligonucleotide (ASO) targeting G4C2 transcripts will allow for the identification of protein signatures associated with a disease-modifying drug. In Specific Aim 3, proteomic signatures common to both human and mouse disease will be targeted in CSF samples from c9ALS patients and asymptomatic C9orf72 expansion carriers, both in cross sectional and longitudinal analyses. The ultimate goal of this project is the discovery of novel biomarkers of disease pathways and progression that will be important for ongoing research into disease pathogenesis and also for future clinical trials.